Microbial biodegradation of 4-chlorobiphenyl, a model compound of chlorinated biphenyls

The biodegradation products of 4-chlorobiphenyl were analyzed in an Achromobacter sp. strain and a Bacillus brevis strain. Both strains generated the same metabolites, with 4-chlorobenzoic acid as the major metabolic product. Our results corroborate previous observations whereby most bacterial strains degrade the chlorobiphenyls via a major pathway which proceeds by an hydroxylation in position 2,3 and a meta-1,2 fission. However, we also detected several metabolites whose structure suggests the existence of other routes for the degradation of chlorinated biphenyls.     

Microbial biodegradation of 4-chlorobiphenyl, a model compound of chlorinated biphenyls.

The biodegradation products of 4-chlorobiphenyl were analyzed in an Achromobacter sp. strain and a Bacillus brevis strain. Both strains generated the same metabolites, with 4-chlorobenzoic acid as the major metabolic product. Our results corroborate previous observations whereby most bacterial strains degrade the chlorobiphenyls via a major pathway which proceeds by an hydroxylation in position 2,3 and a meta-1,2 fission. However, we also detected several metabolites whose structure suggests the existence of other routes for the degradation of chlorinated biphenyls.     

Effect of chlorobenzoates on the degradation of polychlorinated biphenyls (PCB) by Pseudomonas stutzeri

Chlorobenzoic acids (CBA) are frequently dead-end products of partial aerobic biodegradation of polychlorinated biphenyls (PCB). When CBA produced from PCB accumulate in the growth medium, they can inhibit the bacterial growth and consequently, slow down PCB biodegradation. In this study, the effects of seven mono- and dichlorinated CBA on growth of Pseudomonas stutzeri on different substrates and on the PCB degradation by this strain in a liquid mineral medium were tested. 3-CBA was the strongest growth inhibitor for P. stutzeri growing on glucose, benzoate and biphenyl. It was found to inhibit heavily the elimination of some di- and trichlorinated biphenyls. In contrast, its influence on the elimination of more chlorinated congeners was much less significant.The authors are with the Department of Biochemical Technology, Faculty of Chemical Technology, Slovak Technical University, 812 37 Bratislava, Slovakia.     

Isolation and characterization of bacteria degrading polychlorinated biphenyls from transformer oil

Polychlorinated biphenyls from transformer oil were degraded in liquid culture under aerobic conditions using a mixed bacterial culture isolated from a transformer oil sample with a high content of polychlorinated biphenyls and other hydrocarbons. Four strains were identified, three of them corresponded to genusBacillus, the other one toErwinia. Bacteria in the transformer oil could remove as much as 65% of polychlorinated biphenyls (88%W/V in the transformer oil). Additional data showed that the two isolated strains ofB. lentus were able to grow on transformer oil and degrade polychlorinated biphenyls by 80 and 83%. Our results provide evidence that microorganisms occurring in transformer oil have the potential to degrade polychlorinated biphenyls.     

Degradation and dechlorination of low-chlorinated biphenyls by a three-membered bacterial co-culture

A Pseudomonas sp. strain, designated CPE1, was found to be capable of completely mineralizing 4-chlorobiphenyl via 4-chlorobenzoate and of partially dechlorinating 3,4-dichlorobiphenyl in the presence of biphenyl. A three-membered bacterial consortium, designated ECO3, prepared by combining CPE1 with two chlorobenzoate (CBA)-degrading strains, was capable of extensively degrading and dechlorinating all the monochlorinated biphenyls and several dichlorinated biphenyls in the presence of bipheny. Both CPE1 and ECO3 were capable of co-metabolizing several low-chlorinated biphenyl congeners of Fenclor 42 in the presence of biphenyl; however, only in ECO3 cultures were high degradation rates and chloride release observed. The higher rate of degradation and mineralization of some polychlorinated biphenyls (PCBs) of Fenclor 42 due to the concerted action of ECO3 members both on PCBs and CBAs suggested that the removal of CBAs from the culture medium may favour PCB degradation, and, therefore, that CBAs may be ivollved in the regulation of the degradation process of several chlorinated biphenyl congeners.Correspoeence to: F. Fava     

Degradation of Aroclor 1221 and survival of strains in soil microcosms

Three bacterial strains able to use different aromatic compounds as the sole carbon and energy source were tested for their potential to degrade Aroclor 1221 in soil microcosms when present in mixed culture. Disappearance of polychlorinated biphenyls (PCBs), occurrence of metabolites, release of chloride, and survival of the laboratory-selected strains were investigated under different conditions. In principle, complete mineralization of various congeners of Aroclor 1221, a technical mixture of PCBs, by the mixed culture was possible. The autochthonous microflora negatively affected the degradation due to formation of a toxic compound from 4-chlorobenzoate. 4-Chlorobenzoate was produced by one of the added strains, Pseudomonas sp. JHK, during degradation of 4-chlorobiphenyl. The unknown metabolite of 4-chlorobenzoate led to a rapid decrease in viable counts of the laboratory-selected strains in the soil microcosm.Correspondence to: J. Havel     

Biodegradation studies of polyaromatic hydrocarbons in aqueous media

Sixteen bacterial strains isolated from an activated sludge and Mycobacterium ssp. PYR-1 were tested for their ability to degrade polyaromatic hydrocarbons (PAHs). The bacterial strains Pasteurella ssp. (B-2) and Mycobacterium ssp. PYR-1 (AM) showed a high biodegradation potential of three- and four-ring PAHs. Bacterial strain AM was able to degrade up to 80% of three and four-ring PAHs (phenanthrene, fluoranthene and pyrene) within the first month of incubation, while the bacterial strain B-2 achieved the same biodegradation in 2 months. The metabolic pathway of PAH degradation was studied using fluoranthene and the bacterial strain AM. Ninety per cent of fluoranthene was biodegraded within the first 9 d of incubation when applied as a single substrate. Retention factor values from thin-layer chromatography studies, gas chromatography with mass selective detection and tandem mass spectrometry identified 9-fluorenone-1-carboxylic acid as one of the stable metabolic products and from this a fluoranthene biodegradation pathway is proposed.     

A new aerobic gram-positive bacterium with a unique ability to degrade ortho- and para-chlorinated biphenyls

Strain B51 capable of degrading polychlorinated biphenyls (PCB) was isolated from soil contaminated with wastes from the chemical industry. Based on its morphological and chemotaxonomic characteristics, the strain was identified as a Microbacterium sp. Experiments with washed cells showed that strain B51 is able to degrade ortho- and para-substituted mono-, di-, and trichlorinated biphenyls (MCB, DCB, and TCB, respectively). Unlike the known PCB degraders, Microbacterium sp. B51 is able to oxidize the ortho-chlorinated ring of 2,2'-DCB and 2,4'-DCB and the para-chlorinated ring of 4.4'-DCB. The degradation of 2,4'-DCB and 4,4'-DCB was associated with the accumulation of 4-chlorobenzoic acid (4-CBA) in the medium in amounts comprising 80-90% of the theoretical yield. The strain was able to utilize 2-MCB, 2,2'-DCB, and their intermediate 2-CBA and to oxidize the mono(ortho)-chlorinated ring of 2,4,2'-TCB and the di(ortho-para)-chlorinated ring of 2,4,4'-TCB. A mixed culture of Microbacterium sp. B51 and the 4-CBA-degrading bacterium Arthrobacter sp. H15 was found to grow well on 1 g/l 2,4'-DCB as the sole source of carbon and energy.     

Biodegradation of Maya crude oil fractions by bacterial strains and a defined mixed culture isolated from Cyperus laxus rhizosphere soil in a contaminated site

Ten bacterial strains were isolated by enrichment culture, using as carbon sources either aliphatics or an aromatic-polar mixture. Oxygen uptake rate was used as a criterion to determine culture transfer timing at each enrichment stage. Biodegradation of aliphatics (10,000 mg L(-1)) and an aromatic-polar mixture (5000 mg L(-1), 2:1) was evaluated for each of the bacterial strains and for a defined culture made up with a standardized mixture of the isolated strains. Degradation of total hydrocarbons (10,000 mg L(-1)) was also determined for the defined mixed culture. Five bacterial strains were able to degrade more than 50% of the aliphatic fraction. The most extensive biodegradation (74%) was obtained with strain Bs 9A, while strains Ps 2AP and UAM 10AP were able to degrade up to 15% of the aromatic-polar mixture. The defined mixed culture degraded 47% of the aliphatics and 6% of the aromatic-polar mixture. The defined mixed culture was able to degrade about 40% of the aliphatic fraction and 26% of the aromatic fraction when grown in the presence of total hydrocarbons, while these microorganisms did not consume the polar hydrocarbons fraction. The proposed strategy that combines enrichment culture together with oxygen uptake rate allowed the isolation of bacterial strains that are able to degrade specific hydrocarbons fractions at high consumption rates.     

多氯联苯降解菌Burkholderia xenovorans LB400研究进展

Burkholderia xenovorans LB400是一株多氯联苯(polychlorinated biphenyls,PCBs)降解菌,可以氧化含有1?6个氯取代基的多氯联苯。近年来,由于其广泛的底物谱和优异的降解性能,菌株LB400已成为研究原核生物降解多氯联苯的生物化学和分子生物学方面的模式生物。目前关于PCBs的微生物降解研究已不再局限于对微生物资源的挖掘,而是更多地聚焦在LB400等降解菌的PCBs降解基因、降解酶的酶学特性以及酶的人工分子进化等方面。同时,LB400作为早期发现的降解菌,其对多氯联苯的降解途径、底物范围及相关机制也被广泛探讨;但是对于PCBs降解相关基因的调控研究较少。因此,本文以Burkholderia xenovorans LB400对多氯联苯降解为核心,通过综述其代谢途径、代谢相关基因和酶系以及降解应用等方面的研究进展,以期为深入探讨Burkholderia xenovorans LB400的应用以及进一步在遗传、分子和生化水平研究其他多氯联苯降解菌株提供借鉴。     

Evidence for 4-chlorobenzoic acid dehalogenation mediated by plasmids related to pSS50.

The biodegradation of 4-chlorobiphenyl usually proceeds through the intermediate 4-chlorobenzoate. Few bacterial strains can degrade 4-chlorobiphenyl to 4-chlorobenzoate and 4-chlorobenzoate to CO2. This study demonstrates that the 4-chlorobiphenyl-degrading Alcaligenes sp. strain ALP83 can degrade 4-chlorobenzoate to 4-hydroxybenzoate. The dehalogenase activity is correlated with a 10-kb fragment carried on plasmid pSS70.     

Evidence for 4-chlorobenzoic acid dehalogenation mediated by plasmids related to pSS50.

The biodegradation of 4-chlorobiphenyl usually proceeds through the intermediate 4-chlorobenzoate. Few bacterial strains can degrade 4-chlorobiphenyl to 4-chlorobenzoate and 4-chlorobenzoate to CO2. This study demonstrates that the 4-chlorobiphenyl-degrading Alcaligenes sp. strain ALP83 can degrade 4-chlorobenzoate to 4-hydroxybenzoate. The dehalogenase activity is correlated with a 10-kb fragment carried on plasmid pSS70.     

Isolation and Characterization of a Mixed Culture That Degrades Polychlorinated Biphenyls

A mixed culture composed of two Pseudomonas strains, designated as KKL101 and KKS102, was isolated from soil. This mixed culture had an enhanced ability to degrade various polychlorinated biphenyls (PCBs) which include highly chlorinated components. They did not grow individually on the mineral salts medium supplemented with a highly chlorinated PCB (PCB48, a mixture of mainly tetrachlorobiphenyl) and biphenyl. When the spent medium of KKL101 was added to the washed cell preparation of KKS102, however, the latter grew on these carbon sources, producing yellow compounds which were identified as metabolic intermediates of the carbon sources, biphenyl and PCBs. These results suggest that KKL101 produces a growth factor(s) essential for KKS102 to grow on PCBs and that the growth of KKL101 is supported by the metabolic intermediates produced by KKS102. It appears that these two bacterial strains have a symbiotic relationship. From the analysis of the degradation products of various PCB congeners, it was found that strain KKS102 degrades a wide range of PCBs which have been considered to be refractory to biological degradation.     

Metabolism of chlorinated guaiacols by a guaiacol-degrading Acinetobacter junii strain.

The metabolism of chlorinated guaiacols by a pure bacterial strain identified by its ability to use guaiacol as the sole carbon and energy source was studied. This strain, identified as Acinetobacter junii 5ga, was unable to grow on several chlorinated guaiacols and catechols. However, strain 5ga grown on guaiacol degraded 4- and 5-chloroguaiacol and 4,5-dichloroguaiacol. Under the same conditions, these cells did not degrade 6-chloroguaiacol, 4,6-dichloroguaiacol, 4,5,6-trichloroguaiacol, or tetrachloroguaiacol, suggesting that the substitution at the 6 position in the ring prevents metabolism of the compound. Degradation of 4-chloroguaiacol was dependent on the initial ratio between the chlorinated compound and viable cells. Transient formation of chlorocatechols resulting from incubation of cells with 4-chloroguaiacol or 4,5-dichloroguaiacol was suggested by UV spectroscopy. Gas chromatography analyses of samples from cultures of strain 5ga grown on guaiacol and incubated with 4- and 4,5-dichloroguaiacol confirmed the presence of 4-chlorocatechol and 4,5-dichlorocatechol, respectively. The formation of the latter was corroborated by gas chromatography-mass spectrometry. Thus, this strain is able to initiate metabolism of specific chlorinated guaiacols by O-demethylation. The starting chlorinated guaiacols and their O-demethylated metabolites inhibited the growth of A. junii 5ga on guaiacol.     

Biodegradation of chlorinated ethenes by a methane-utilizing mixed culture

Chlorinated ethenes are toxic substances which are widely distributed groundwater contaminants and are persistent in the subsurface environment. Reports on the biodegradation of these compounds under anaerobic conditions which might occur naturally in groundwater show that these substances degrade very slowly, if at all. Previous attempts to degrade chlorinated ethenes aerobically have produced conflicting results. A mixed culture containing methane-utilizing bacteria was obtained by methane enrichment of a sediment sample. Biodegradation experiments carried out in sealed culture bottles with radioactively labeled trichloroethylene (TCE) showed that approximately half of the radioactive carbon had been converted to 14CO2 and bacterial biomass. In addition to TCE, vinyl chloride and vinylidene chloride could be degraded to products which are not volatile chlorinated substances and are therefore likely to be further degraded to CO2. Two other chlorinated ethenes, cis and trans-1,2-dichloroethylene, were shown to degrade to chlorinated products, which appeared to degrade further. A sixth chlorinated ethene, tetrachloroethylene, was not degraded by the methane-utilizing culture under these conditions. The biodegradation of TCE was inhibited by acetylene, a specific inhibitor of methane oxidation by methanotrophs. This observation supported the hypothesis that a methanotroph is responsible for the observed biodegradations.     

Biodegradation of chlorinated ethenes by a methane-utilizing mixed culture.

Chlorinated ethenes are toxic substances which are widely distributed groundwater contaminants and are persistent in the subsurface environment. Reports on the biodegradation of these compounds under anaerobic conditions which might occur naturally in groundwater show that these substances degrade very slowly, if at all. Previous attempts to degrade chlorinated ethenes aerobically have produced conflicting results. A mixed culture containing methane-utilizing bacteria was obtained by methane enrichment of a sediment sample. Biodegradation experiments carried out in sealed culture bottles with radioactively labeled trichloroethylene (TCE) showed that approximately half of the radioactive carbon had been converted to 14CO2 and bacterial biomass. In addition to TCE, vinyl chloride and vinylidene chloride could be degraded to products which are not volatile chlorinated substances and are therefore likely to be further degraded to CO2. Two other chlorinated ethenes, cis and trans-1,2-dichloroethylene, were shown to degrade to chlorinated products, which appeared to degrade further. A sixth chlorinated ethene, tetrachloroethylene, was not degraded by the methane-utilizing culture under these conditions. The biodegradation of TCE was inhibited by acetylene, a specific inhibitor of methane oxidation by methanotrophs. This observation supported the hypothesis that a methanotroph is responsible for the observed biodegradations.     

好氧氯苯降解菌的分离鉴定

【目的】分离好氧氯苯降解菌,并通过研究降解特性为应用提供理论依据。【方法】利用富集培养技术分离菌株,通过形态、生理生化反应特征及16S rRNA基因序列分析鉴定菌株,测定培养液中氯苯、其它氯苯类化合物和氯离子的浓度以及菌体细胞的密度和菌体细胞粗提液中邻苯二酚双加氧酶的活性,研究菌株的降解特性。【结果】16S rRNA基因序列相似性比较表明,分离出的菌株与乙酸钙不动杆菌(Acinetobacter calcoaceticus)的相似性高达98.5%。以初始浓度为50mg/L的氯苯为唯一碳源和能源时,120h内菌株对氯苯的降解率高达98.2%,氯离子净释放量和氯苯降解量的摩尔比范围为1:1.85-1:1.39,菌体细胞粗提液中邻苯二酚1,2-双加氧酶的平均活性为0.538U/mg蛋白质。加入葡萄糖后,菌体细胞数量和氯离子浓度明显增加,但单位细胞的氯苯降解能力明显下降。在二氯苯和三氯苯共存时,菌株对氯苯的降解能力受到明显的抑制作用,但对二氯苯有一定的降解作用,降解能力大小顺序为:1,3-二氯苯1,2-二氯苯1,4-二氯苯。【结论】分离出的好氧氯苯降解菌属于Acinetobacter属菌株,该菌株对氯苯和二氯苯均具有降解作用,可能通过邻位裂环途径降解氯苯,氯苯对菌株的降解能力和邻苯二酚1,2-双加氧酶的活性具有明显的增强作用。     

Degradation of Microcystin-RR by Sphingomonas sp. CBA4 isolated from San Roque reservoir (Córdoba – Argentina)

We report the aerobic biodegradation of Microcystin-RR (MC-RR) by a bacterial strain isolated from San Roque reservoir (Córdoba – Argentina). This bacterium was identified as Sphingomonas sp. (CBA4) on the basis of 16S rDNA sequencing. The isolated strain was capable of degrading completely MC-RR (200 μg l⁻¹) within 36 h. We have found evidence that MC-RR biodegradation pathway by this Sphingomonas sp. strain would start by demethylating MC-RR, affording an intermediate product, which is finally biodegraded by this strain within 72 h. Our results confirm that certain environmental bacteria, living in the same habitat as toxic cyanobacteria, have the capability to perform complete biodegradation of MC, leading to natural bioremediation of waterbodies. The bacterium reported here presents genetic homologies with other strains that degrade MC-LR. However, initial demethylation of MC-RR has been not described previously, raising questions on the probable presence of different biodegradation pathways for different MC variants.     

Total biodegradation of the oestrogenic mycotoxin zearalenone by a bacterial culture

A mixed culture of bacteria, enriched from soil collected at a coal gasification site, proved capable of removing the potent oestrogenic mycotoxin zearalenone from culture media. The bacteria grew rapidly when zearalenone was provided as the sole source of carbon and energy. HPLC and ELISA analysis of culture extracts revealed no zearalenone or zearalenone-like products. Fourteen bacterial isolates from the mixed culture were identified and purified. The ability to degrade zearalenone was lost upon purification and recombination of the bacterial members of the mixed culture. A strain of Pseudomonas fluorescens capable of degrading polychlorinated biphenyls was unable to degrade zearalenone. This is the first report of the complete degradation of zearalenone by bacteria. The present study suggests the potential of mixed cultures in the biodegradation of zearalenone.     

Catabolic degradation of 4-chlorobiphenyl byPseudomonas sp. DJ-12 via consecutive reaction ofmeta-cleavage and hydrolytic dechlorination

Pseudomonas sp. strain DJ-12 is a bacterial isolate capable of degrading 4-chlorobiphenyl (4CBP) as a carbon and energy source. The catabolic degradation of 4CBP by the strain DJ-12 was studied along with the genetic organization of the genes responsible for the crucial steps of the catabolic degradation. The catabolic pathway was characterized as being conducted by consecutive reactions of themeta-cleavage of 4CBP, hydrolytic dechlorination of 4-chlorobenzoate (4CBA), hydroxylation of 4-hydroxybenzoate, andmeta-cleavage of protocatechuate. ThepcbC gene responsible for themeta-cleavage of 4CBP only showed a 30 to 40% homology in its deduced amino acid sequence compared to those of the corresponding genes from other strains. The amino acid sequence of 4CBA-CoA dechlorinase showed an 86% homology with that ofPseudomonas sp. CBS3, yet only a 50% homology with that ofArthrobacter spp. However, thefcb genes for the hydrolytic dechlorination of 4CBA inPseudomonas sp. DJ-12 showed an uniquely different organization from those of CBS3 and other reported strains. Accordingly, these results indicate that strain DJ-12 can degrade 4CBP completely viameta-cleavage and hydrolytic dechlorination using enzymes that are uniquely different in their amino acid sequences from those of other bacterial strains with the same degradation activities.